Abstract Triacetic acid lactone (TAL) is a platform chemical biosynthesized primarily through decarboxylative Claisen condensation by type III polyketide synthase 2-pyrone synthase (2-PS). However, this reaction suffers from intrinsic energy inefficiency and feedback inhibition by and competition for malonyl-CoA. TAL production through non-decarboxylative Claisen condensation by polyketoacyl-CoA thiolase alleviates many of these disadvantages. We discovered five more thiolases with TAL production activity by exploring homologs of a previously reported polyketoacyl-CoA thiolase, BktB, from Cupriavidus necator . Among them, the BktB homolog from Burkholderia sp. RF2-non_BP3 has ∼ 30 times higher in vitro and in vivo TAL production activity and led to ∼10 times higher TAL titer than 2-PS when expressed in Escherichia coli , achieving a titer of 2.8 g/L in fed-batch fermentations. This discovery of a novel polyketoacyl-CoA thiolase with superior TAL production activity paves the way for realization of total biomanufacturing of TAL.

A highly activeBurkholderiapolyketoacyl-CoA thiolase for production of triacetic acid lactone